LOCUS       sequence-476949-        6220 bp    DNA     circular SYN 24-JUN-2025
DEFINITION  Expresses 3XFLAG::GR LBD::DAF-16A from the hsp16.48 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6220)
  AUTHORS   Monsalve GC, Yamamoto KR, Ward JD
  TITLE     A New Tool for Inducible Gene Expression in Caenorhabditis elegans.
  JOURNAL   Genetics. 2019 Feb;211(2):419-430. doi: 10.1534/genetics.118.301705.
            Epub 2018 Nov 30.
  PUBMED    30504365
REFERENCE   2  (bases 1 to 6220)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jun 24, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1534/genetics.118.301705"; journalName: "Genetics"; date: 
            "2019-02"; volume: "211"; issue: "2"; pages: "419-430"
COMMENT     Sequence Label: pGM59
FEATURES             Location/Qualifiers
     source          1..6220
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(33..461)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(143..164)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     355..374
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     588..610
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     602..619
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     603..619
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        730..1034
                     /label=hsp16-48 promoter
                     /note="promoter for the C. elegans heat shock protein 16-48
                     gene (Russnak and Candido, 1985)"
     CDS             1048..1113
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1120..1140
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     terminator      3604..3927
                     /gene="tbb-2"
                     /label=tbb-2 terminator
                     /note="transcription terminator for the C. elegans tubulin 
                     beta-2 gene"
     promoter        complement(4035..4053)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4074..4090)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4074..4090)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4087..4109)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4098..4114
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4122..4128,4129..4146,4147..4152))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4167..4188
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4305..4322)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4476..5064)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4556..4575)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5235..6026,6027..6095))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5858..5877
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6096..6200)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
       61 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
      121 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
      181 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
      241 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
      301 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
      361 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
      421 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct
      481 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
      541 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
      601 ttgtaaaacg acggccagtg aattgtaata cgactcacta tagggcgaat tgggtaccgc
      661 tggacggaaa tagtggtaaa gtgacatgat tgtagtttga agatttcaca attagagtga
      721 atgttgtttg gttcggtttt gtcactgtat ttatactcat ttccaccttt ttctagaaca
      781 ttcgagctgc ttcttacaaa aggaggcgac tcacattcag aacattgaga aatagtgtgc
      841 gactgaagaa acccagatac tttttcaatc tgcgtctctt tgcacctatg gggtgtattt
      901 tgaaatgaat gcatctagga ccttctagaa cattctaaac ggctgcagga tacgggtata
      961 taagccaatc gtgttcagag gaaaccaata cactttgttc aagtgcttac tgttcattct
     1021 ctaaacttca agaactagaa aaaaatggac tacaaggacc acgacggaga ctacaaggac
     1081 cacgacatcg actacaagga cgacgacgac aagatggccc caaagaagaa gcgtaaggtc
     1141 ggaatccacg gagtcccagc cgccattcag caggccacta caggagtctc acaagaaacc
     1201 tctgaaaatc ctggtaacaa aacaatagta cctgcaacgt taccacaact cacccctacc
     1261 ctggtgtcac tgttggaggt tattgaacct gaagtgttat atgcaggata tgatagctct
     1321 gttccagact caacttggag gatcatgact acgctcaaca tgttaggagg gcggcaagtg
     1381 attgcagcag tgaaatgggc aaaggcaata ccaggtttca ggaacttaca cctggatgac
     1441 caaatgaccc tactgcagta ctcctggatg tttcttatgg catttgctct ggggtggaga
     1501 tcatatagac aatcaagtgc aaacctgctg tgttttgctc ctgatctgat tattaatgag
     1561 cagagaatga ctctaccctg catgtacgac caatgtaaac acatgctgta tgtttcctct
     1621 gagttacaca ggcttcaggt atcttatgaa gagtatctct gtatgaaaac cttactgctt
     1681 ctctcttcag ttcctaagga cggtctgaag agccaagagc tatttgatga aattagaatg
     1741 acctacatca aagagctagg aaaagccatt gtcaagaggg aaggaaactc cagccagaac
     1801 tggcagcggt tttatcaact gacaaaactc ttggattcta tgcatgaagt ggttgaaaat
     1861 ctccttaact attgcttcca aacatttttg gataagacca tgagtattga attccccgag
     1921 atgttagctg aaatcatcac caatcagata ccaaaatatt caaatggaaa tatcaaaaaa
     1981 cttctgtttc atcaaaagct gcagatgaac gactcaatag acgacgattt tccgccggag
     2041 ccacgtggca ggtgttacac gtggccaatg caacaataca tctatcagga atcgtcagca
     2101 accattcccc atcaccattt aaatcaacac aacaatccgt atcatccaat gcatcctcat
     2161 catcaattac ctcatatgca acaacttcct caacctctat tgaatcttaa catgacgacg
     2221 ttaacatctt ctggcagttc cgtggccagt tccattggag gcggagctca atgctctccg
     2281 tgcgcgtcgg gctcctcgac cgctgcaaca aattcctctc aacagcagca gaccgttggt
     2341 caaatgcttg ctgcatcggt gccttgttct tcatctggca tgacacttgg aatgtcactt
     2401 aatctgtcac aaggcggtgg tccaatgccg gcaaaaaaga agcgttgtcg taagaagcca
     2461 accgatcaat tggcacagaa gaaaccgaat ccatggggtg aggaatccta ttcggatatc
     2521 attgccaaag cattggaatc ggcgccagac ggaaggctta aactcaatga gatttatcaa
     2581 tggttctctg ataatattcc ctactttgga gaacgatcta gtcccgagga ggccgccgga
     2641 tggaagaact cgatccgtca caatctgtct cttcattctc gtttcatgcg aattcagaat
     2701 gaaggagccg gaaagagctc gtggtgggtt attaatccag atgcaaagcc aggaaggaat
     2761 ccacggcgta cacgtgaacg atccaatact attgagacga ctacaaaggc tcaactcgaa
     2821 aaatctcgcc gcggagccaa gaagaggata aaggagagag cattgatggg ctcccttcac
     2881 tcgacactta atggaaattc gattgccgga tcgattcaaa cgatttctca cgatttgtat
     2941 gatgatgatt caatgcaagg agcatttgat aacgttccat catctttccg tccccgaact
     3001 caatcgaacc tctcgattcc tggatcgtcg tctcgtgttt ctccagctat tggaagtgat
     3061 atctatgatg atctagaatt cccatcatgg gttggcgaat cggttccagc aattccaagt
     3121 gatattgttg atagaactga tcaaatgcgt atcgatgcaa ctactcatat tggtggagtt
     3181 cagattaagc aggagtcgaa gccgattaag acggaaccaa ttgctccacc accatcatac
     3241 cacgagttga acagtgtccg tggatcgtgt gctcagaatc cacttcttcg aaatccaatt
     3301 gtgccaagca ctaacttcaa gccaatgcca ctaccgggtg cctatggaaa ctatcaaaat
     3361 ggtggaataa ctccaatcaa ttggctatca acatccaact catctccact gcctggaatt
     3421 caatcgtgtg gaattgtagc tgcacagcat actgtcgctt cttcatcggc tcttccaatt
     3481 gatttggaaa atctgacact tcccgatcag ccactgatgg atactatgga tgttgatgca
     3541 ttgatcagac atgagctgag tcaagctgga gggcagcata ttcattttga tttgtaagaa
     3601 ttcatgcaag atcctttcaa gcattccctt cttctctatc actcttcttt ctttttgtca
     3661 aaaaattctc tcgctaattt atttgctttt ttaatgttat tattttatga ctttttatag
     3721 tcactgaaaa gtttgcatct gagtgaagtg aatgctatca aaatgtgatt ctgtctgatg
     3781 tactttcaca atctctcttc aattccattt tgaagtgctt taaacccgaa aggttgagaa
     3841 aaatgcgagc gctcaaatat ttgtattgtg ttcgttgagt gacccaacaa aaagaggaaa
     3901 ctttattgtg ccgccaagaa aaaagtctca ttgcctaaga gaaacatatg tggttggaaa
     3961 tcttccagat cgtggatcac tagtatgtgg ttggaaatct tccagatcgt ggatcagagc
     4021 tccagctttt gttcccttta gtgagggtta atttcgagct tggcgtaatc atggtcatag
     4081 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     4141 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     4201 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     4261 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     4321 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     4381 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     4441 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     4501 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     4561 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     4621 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     4681 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     4741 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     4801 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     4861 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     4921 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     4981 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     5041 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     5101 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     5161 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     5221 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     5281 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     5341 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     5401 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     5461 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     5521 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     5581 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     5641 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     5701 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     5761 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     5821 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     5881 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     5941 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     6001 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     6061 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     6121 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     6181 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
//
